The development of appropriate neuronal circuitry is critical for normal function of the nervous system. Defects in the formation or modulation of synaptic connections may contribute to the etiology of a number of developmental and nervous system disorders. Yet little is known about the molecules that regulate synapse formation and function. In the peripheral nervous system, nerve growth factor (NGF) has been implicated in the innervation of target tissues by sympathetic neurons. Recent studies in this laboratory have indicated that NGF also influences the formation and strength of synapses between sympathetic neurons and cardiac myocytes in vitro. This system provides a novel opportunity to investigate both developmental and modulatory roles of neurotrophins in the establishment of neuronal circuitry. The goals of this proposal are to define the cellular and molecular mechanisms underlying NGF-mediated enhancements in synapse formation and function. Synaptic connections that form between sympathetic neurons and cardiac myocytes can be assayed physiologically by measuring evoked postsynaptic currents in myocytes, or functionally as an increase in myocyte beat rate upon stimulation of a connected neuron. The neurotransmitter dependence, neurotrophin NGF-dependent synaptic modulation will be determined in neuron/myocyte co-cultures. We will determine whether NGF acts presynaptically to alter neurotransmitter release or postsynaptically to alter the myocyte response to neurotransmitter. We will test the hypothesis that NGF acutely potentiates synaptic transmission by enhancing the release of norepinephrine from presynaptic terminals and will investigate a role for additional co-transmitters in the system. Preliminary results indicate that NGF-dependent modulation of sympathetic synapses may involve two different receptors for NGF, TrkA and p75. We will investigate whether the level of NGF-dependent synaptic modulation is regulated through two interacting signal transduction systems. Finally, we have demonstrated long-term effects of NGF on the development of sympathetic synapses. We will investigate whether NGF determines the number or strength of synapses that develop between sympathetic neurons and cardiac myocytes. These experiments will, for the first time, define the developmental and modulatory roles of NGF at the synaptic level.